Abstract Current influenza (flu) vaccines are effective only for closely matched flu viral strains and must be updated annually to address constant antigenic shift/drifts of surface hemagglutinin (HA) and/or neuraminidase (NA) of the virus. Even with annual update, there have been years in which flu vaccines were ineffective due to significant differences in antigenicity of HA and/or NA between the strains used for preparing the vaccines and the circulating ones, leaving us at high risk of pandemics in case a new and highly pathological virus emerges. It is without any doubt that a ?Universal? flu vaccine that can protect against both seasonal (matched or mismatched) and pandemic flu viruses is urgently needed, but it remains ?an alchemist?s dream? so far. We developed a novel adjuvant by encapsulation of cGAMP, an agonist of the stimulator of interferon (IFN) gene (STING), into pulmonary surfactant (PS)-biomimetic liposomes (PS-GMNP). The adjuvant, alongside an inactivated flu vaccine, robustly stimulated humoral and CD8+ T cell immune responses that resemble those ocurring during the early phase of viral infection both in magnitude and in dynamics. Strikingly, a single dose of PS-GMNP-adjuvant flu vaccine elicited strong cross-protection against a lethal challenge of diverse heterosubtypic flu A viruses as early as 2 days after immunization. While stimulating robust heterosubtypic immunity, the adjuvant did not cause any adverse events in lung histology, body weight or temperature, in sharp contrast to the severe lung inflammation and death caused by flu viral infection. In this proposal, we will investigate the cellular and humoral immune responses essential to the cross-protection induced by PS- GMNP-adjuvanted flu vaccines. Specifically, we will determine whether PS-GMNP can expand cross-reactive CD8+ T cells and induce broadly neutralizing antibodies (bnAbs), pivotal to PS-GMNP-induced cross- protection. In Aim 2, we will preclude any adverse effects of pre-existing immunity on the adjuvanticity of PS- GMNP and extend the cross-protection to pre-pandemic wild type H5N1 and H7N9 viruses to establish its clinical potentials. Distinguished from conventional adjuvants that activate primarily antigen-presenting cells (APCs), cGAMP delivered by PS-GMNP activated both alveolar macrophages (aM?) and alveolar epithelial cells (AEC), which can be crucial since similar activation of these two types of cells is also observed during flu viral infection. Two alternative approaches will be employed in Aim 3 to corroborate indispensable function of AEC and/or aM? in PS-GMNP-mediated heterosubtypic immunity, including a blockade of cell-gap junctions and generation of chimeric mice of wild type and STING-deficient bone marrow (BM) cells. The study, if successful, could provide invaluable information about clinical potentials for PS-GMNP to widen the breadth of existing flu vaccines toward a ?Universe? one, which will have a huge and immediate impact on global health.